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An optimized method for mouse liver sinusoidal endothelial cell isolation.
- Source :
-
Experimental Cell Research . Dec2016, Vol. 349 Issue 2, p291-301. 11p. - Publication Year :
- 2016
-
Abstract
- The objective of the present study was to develop an accurate and reproducible method for liver sinusoidal endothelial cell (LSEC) isolation in mice. Non-parenchymal cells were isolated using a modified two-step collagenase digestion combined with Optiprep density gradient centrifugation. LSEC were further purified using two prevalent methods, short-term selective adherence and CD146+ magnetic-activated cell sorting (MACS), and compared in terms of cell yield, viability and purity to our purification technique using CD11b cell depletion combined with long-term selective adherence. LSEC purification using our technique allowed to obtain 7.07±3.80 million LSEC per liver, while CD146+ MACS and short-term selective adherence yielded 2.94±1.28 and 0.99±0.66 million LSEC, respectively. Purity of the final cell preparation reached 95.10±2.58% when using our method. In contrast, CD146+ MACS and short-term selective adherence gave purities of 86.75±3.26% and 47.95±9.82%, respectively. Similarly, contamination by non-LSEC was the lowest when purification was performed using our technique, with a proportion of contaminating macrophages of only 1.87±0.77%. Further, isolated cells analysed by scanning electron microscopy presented typical LSEC fenestrations organized in sieve plates, demonstrating that the technique allowed to isolate bona fide LSEC. In conclusion, we described a reliable and reproducible technique for the isolation of high yields of pure LSEC in mice. This protocol provides an efficient method to prepare LSEC for studying their biological functions. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 00144827
- Volume :
- 349
- Issue :
- 2
- Database :
- Academic Search Index
- Journal :
- Experimental Cell Research
- Publication Type :
- Academic Journal
- Accession number :
- 119581619
- Full Text :
- https://doi.org/10.1016/j.yexcr.2016.10.024